Telomeres are DNA-protein complexes that play important roles in the replication and protection of the chromosome ends. Telomere maintenance is essential for genomic stability, and telomeric DNA loss associated with replication triggers cellular senescence. For tumor cells, maintaining stable telomere length is required for sustained growth. In human, several protein factors are involved in regulating telomere length. TRF1 is a telomeric DNA binding protein that negatively influences telomere length homeostasis. Tankyrase1 and TIN2 are tethered to telomeres through interacting with TRF1. Both tankyrase1 and TIN2 are implicated in human telomere length regulation. Tankyrase1 is a telomeric poly(ADP-ribose) polymerase that adds poly(ADP-ribose) side chain to TRF1 molecule, removing it from telomeres. When over-expressed, tankyrase1 promotes telomere elongation, an effect opposite to that of TRF1. Over-expression of truncated TIN2 causes telomere elongation. Since members of the TRF1 protein complex play active roles in telomere length homeostasis in human cells, they may provide therapeutic targets for cancer therapy. Characterizing the interplay among members of TRF1 protein complex is critical for designing treatment approaches which target the TRF1 protein complex. TIN2 may function as a tankyrase1 inhibitor in TRF1 complex. We will test the potential role of TIN2's in regulating the functional status of the TRF1 complex, with regard to telomere length maintenance in human tumor cells. These experiments are designed to test the biological basis for developing telomere-based treatment protocols. The feasibility of targeting tankyrase1 for cancer treatment will also be tested in this grant proposal. Since tankyrase1 is a poly(ADP-ribose) polymerase whose enzymatic activity is critical for telomere homeostasis, it is conceivable that specific tankyrase1 inhibitors can be useful in cancer therapy. To test this hypothesis, a dominant-negative tankyrase1 allele will be introduced into human tumor cells and the changes in telomere length and growth potential will be studied. The potential synergy between tankyrase1 inhibition and other cancer treatment modalities (telomerase inhibitors, chemotherapy, and radiation therapy) will also be tested in tissue culture systems.